Program Director/Principal Investigator (Last, First, Middle): Su, Gloria H. PROJECT SUMMARY The majority of human pancreatic cancer is presented as pancreatic ductal adenocarcinoma (PDA). Oncogene Kras and tumor-suppressor gene p16 are the two most frequently mutated genes in PDA. Based on the genetics of PDA, we have developed two mouse models, p16-/-;LSL-KRASG12D;PDX1-Cre mice and p16lox/lox;LSL-KRASG12D;PDX1-Cre mice, which harbor an activated oncogenic Kras and biallelically inactivated p16 in their pancreases. Our data shows that both of these models develop mPanIN (precancerous lesions similar to those observed in humans), invasive cancer (similar to PDA), and metastasis. Here we propose to utilize these mouse models and cancer cell lines derived from the tumors and metastases of these mice to further our understandings in the differential roles of the wild-type Kras and oncogenic Kras in metastasis, and to investigate if the restoration of p16 will be sufficient to stop tumor development in vivo. Our preliminary data showed that in p16-/-;LSL-KRASG12D;PDX1-Cre mice, the wild-type Kras allele was selectively inactivated as tumors progressed to metastases, while the mutant Kras was retained at 100%. This suggests that the wild-type Kras may have a role in inhibiting metastasis and the oncogenic Kras has additional functions beyond tumor initiation and may promote metastasis. In Aim 1, we propose to utilize our unique panel of p16-/-;KRASG12D/+ and p16-/-;KRASG12D/- cancer cell lines to study the functions of the wild-type and mutant Kras in metastasis using in vitro assays, subcutaneous and orthotopic mouse models, and Lentivirus shRNA knockdown experiments. The restoration of p16 can lead to reduced growth rate, increased hypophosphorylated Rb, accumulated cells in G1, induction of senescence, and sometimes apoptosis in vitro. In Aim 2, we propose to investigate the impact of p16 restoration in our unique panel of p16-/-;KRASG12D/+ cancer cell lines as well as in our p16-/-;LSL-KRASG12D;PDX1-Cre mice. We propose to generate a tamoxifen inducible p16 knock-in mouse line (p16KI), which will allow us to restore p16 at will. By breed p16-/-;LSL- KRASG12D;PDX1-Cre mice to the p16KI mice, we will be able to restore p16 at various stages of pancreatic tumorigenesis and assess its impact on in vivo tumor progression. Finally in Aim 3, we propose to continue our characterization of Smad4lox/lox;PDX1-Cre;MT-TGFalpha mice, which we have developed to investigate human PDA that do not harbor Kras mutations. Our preliminary data showed that these mice can develop mPanINs. We propose to investigate the mPanINs, invasive cancer, and metastasis in this model for pathological and genetic changes to further our understanding of human PDA. In summary, we have developed three mouse models that closely simulate human PDA in pathology and genetics. In this funding cycle, we will utilize these models to further our understandings of metastasis and to design better treatment options (specifically targeting Kras and p16). PHS 398/2590 (Rev. 11/07) Page Continuation Format Page